ACTIVATION OF THE NADPH OXIDASE IN HUMAN FIBROBLASTS BY MECHANICAL INTRUSION OF A SINGLE-CELL WITH AN ULTRAMICROELECTRODE

We present here a real-time and single cell study of an oxidative stre ss mechanism in human fibroblasts, Hydrogen peroxide released by a sin gle normal or SV40-transformed human fibroblast was detected at the su rface of an ultramicroelectrode while puncturing the cell membrane wit h the ultramicroelectrode tip itself or,vith a micropipette, This mech anical intrusion induced the emission of large quantities (10(-15)-10( -14) mel) of H2O2 by the cell with a very short time delay (<0.5 s). W e show that this H2O2 production was an active neo-production by fibro blasts when the membrane was stressed by the cellular puncture and is a model which could mimic similar effects as particle (virus, bacteria , etc.) intrusion into the cell, Cell incubations in the presence of s ome inhibitors of the different NADPH oxidase enzymes, using ultramicr oelectrode measurements of the short time effects (<20 min) let us bel ieve that an NADPH oxidase-like enzyme may be implicated in this induc ed-H2O2 generation, Phenylarsine oxide (PAO), a specific NADPH oxidase inhibitor, at concentrations between 0.5-50 mu M seemed to quickly ki ll the transformed cells preferentially to the normal cells, pointing out for the future a possible anti-cancerous chemotherapic use.